The purpose of the proposed project is to determine the biochemical and genetic mechanisms regulating expression of genes during fungal development. The organism to be studied is the genetically well characterized Ascomycete, Aspergillus nidulans. A. nidulans reproduces asexually by forming multicellular conidiophores and uninucleate conidia. During the previous funding period we cloned and characterized numerous genes that are specifically activated during development, including three major regulatory genes. These three genes, designated brlA, abaA, and wetA, will be the focus of our work during the next funding period. The cloned genes will be used in conjunction with identified developmental regulatory genes exert their activities and to identify additional regulatory elements. Five sets of experiments with interrelated objectives are planned. Initially we will complete analysis of brlA, abaA, and wetA by inducing expression of wetA in vegetative cells and by determining if these genes are autoregulated. We will identify and characterize cis-acting regulatory elements adjacent to conidiation-specific genes, including the three regulatory genes. We will test the hypothesis that the regulatory genes directly control the activities of one another and the many genes whose products contribute directly to conidiophore form and function. We will conduct mutational searches for new regulatory genes whose products interact with those of the previously identified regulatory genes. Finally, we will begin characterization of a gene that encodes and abundant, developmentally regulated, basic, nuclear protein that may be involved in the generalized transcriptional shutdown that occurs upon induction of the conidiation pathway. This work will provide important new information about the reproductive molecular biology and genetics of a group of medically, industrially, and agriculturally important organisms. The results may suggest methods and approaches for controlling the beneficial and detrimental activities of these fungi. In addition, they will be relevant to the broader question of how multicellular organisms coordinate the expression of the many genes contributing to the phenotypes of differentiated cells.